Ribosome regulation gone viral!

Welcome to the first post of Tucking Into Science!! There will be many more posts to come in the future but to start off with I’ll be looking at a paper entitled “RNA regulons in Hox 5′ UTRs confer ribosome specificity to gene regulation” by Xue et al.

Ribosomes are complex molecules present in the majority cells, that play a key role in the synthesis of new proteins. They function by binding and moving along a strand of mRNA, whilst ‘reading’ the sequence order of nucleotide bases and attaching corresponding amino acids together to form a protein (see figure below). Protein synthesis is an ongoing critical process throughout the life and growth of an organism.

In the developmental stages of an animal, proteins of homeobox genes (Hox) – genes whose products form the basic body plan of an animal such as skeletal and nervous tissue – are produced at precise points throughout the developmental process. Regulation of ribosome binding to the mRNA at these stages is key for correct translation and protein synthesis. A new method of regulation for this ribosomal binding has recently been found in eukaryotic homeobox genes.

In eukaryotic cells the dominant method of regulation is by the addition of a guanine nucleotide ‘cap’ to the 5’ end of the mRNA, consisting of a methylated guanine nucleotide, linked to the mRNA by a triphosphate linker. In cap-dependent translation (CDT) this cap facilitates the binding of initiation factors, leading to the formation of the preinitiation complex (PIC). The PIC scans the nucleotide strand for the start codon and subsequently recruits the ribosome for translation of the mRNA. In viruses that don’t possess capped mRNA, ribosomes are directly recruited to the mRNA by a specific nucleotide sequence present in the 5’ untranslated region (UTR) called an internal ribosome entry site (IRES). This process is known as IRES-dependent translation (IDT) and provides an alternative route of initiation. It has previously been found that IRESs are present in a small amount of cellular genes and considered to act as a failsafe for conditions where rate of translation is reduced, but full extent of its control is unknown. However, a paper published by Xue et al. in Nature has found evidence supporting the idea that IRESs are present and crucial in mammalian ribosome regulation of Hox genes.

Using multipotent mouse stem cells, Xue and colleagues determined whether cap-independent translation is established through 5’ UTRs. They measured the protein expression levels of two different types of luciferase; one expressed by cap-dependent translations and the other’s whose expression was dependent upon if the 5’ UTRs could recruit ribosomes. Through this it was unexpectedly shown that a number of Hox 5’ UTRs contain IRESs and using further controls composed of single Hox gene transcripts, established a correlation between the presence of IRES elements and regulation by a ribosome protein, RPL38 (especially for the Hox gene subset HoxA). RPL38 is a key protein of the eighty that form the eukaryotic ribosome and previously shown to be a regulator of ribosomes by facilitating its formation. RPL38 was later confirmed having a significant role in IRES-dependent translation regulation by examining the effects of its removal from the murine cells.

The next step of the authors was to investigate the sequence in the Hox 5’ UTR that was crucial for IDT. Through deleting sections of the 1.2 kilobase Hoxa9 5’ UTR, a domain of around 300 bases was determined, having a high evolutionary conservation throughout vertebrates and required for IRES function. Elaborating on the conserved sequence, modelling was used to establish the structure; a four-way junction with additional long arm hairpin arms and a right angle asymmetric bulge. The bulge is not only similar between cellular HoxA genes but also bears structural resemblance to that of the IRES in the Hepatitis C virus, which has led to the suggestion that certain structures in the IRES element are required for activity. This structure potentially interacts with RPL38 to recruit the ribosome directly to the mRNA (see figure below).

Moreover, it was found that the removal of either or both of the hairpins resulted in a dramatic IRES activity decrease. Once Xue et al. had established the presence and general structure of the IRES elements in Hox 5’ UTRs, they looked at the role of these element in normal translation. By removing the IRESs from the UTR, it was found that translation noticeably decreased even though the mRNA remained capped, leading to the idea that some additional elements in the 5’ UTR inhibit CDT. In the gene Hoxa9 a domain in the first 300 nucleotides is solely able to strongly reduce levels of CDT, termed a translational inhibitory element (TIE). TIEs have been postulated to obstruct the movement of the PIC in the 3’ direction (see figure above). These elements allow the Hox mRNAs to be regulated independently from the other mRNAs in the cell by seemingly blocking the PIC’s progress and preventing CDT.

Xue et al. have highlighted the previously unknown critical function in Hox gene translation and thus in mammalian development. This discovery could lead onto advances in treatments for diseases that target ribosomal regulation. This could include being able to reduce the amount of amyloid precursor protein in the brain which can lead to Alzheimer’s disease. However further research is needed to establish the complete role IRESs have in ribosomal regulation and the full extent of their presence in eukaryotic cells.

If you would like to read the paper itself, just follow the link below: